Electrophysiological characterization of a tobacco
cyclic nucleotide-gated channel (“NtCBP4”) expressed in oocytes.
All recordings were made in the two-electrode configuration, and are
presented without leak subtraction. Command voltages were applied in
20-mV steps between −60 and −160 mV. In all cases, [dibutyryl-cAMP]
was 5 mm, and the standard oocyte bath solution was used
(i.e. with 96 mm K+). A,
Current-voltage relationship of NtCBP4 currents. NtCBP4 showed no
(inward) currents before addition of dibutyryl-cAMP to the perfusion
bath solution (▴). After application of dibutyryl cAMP to the
perfusion bath, currents were monitored over 20-min intervals (data not
shown), and reached maximum values by (approximately) 60 min (●) in
this experiment. Water-injected oocytes displayed no inward currents
after exposure to dibutyryl-cAMP for 60 min (○). Results are presented as
means (n = 4) ± se. The
insert shows representative time-dependent currents recorded from
oocytes expressing NtCBP4 before (bottom of insert) and 60 min after
addition of dibutyryl cAMP (top of insert). B, Increase of
cAMP-activated NtCBP4 currents over time. Time-dependent current
recordings are shown for two oocytes expressing NtCBP4. In one case
(cell 1), current is shown to be greater at 60 min than that recorded
after 30-min incubation in perfusion bath solution containing
dibutyryl-cAMP. In the case of the second cell, noticeable currents
were recorded only after 100 min incubation in dibutyryl-cAMP; these
currents were substantially lower than those recorded from cell one,
however. C, Reversal of cAMP activation of plant cngc currents. The top
time-dependent currents were recorded from an oocyte expressing NtCBP4
before addition of dibutyryl-cAMP to the perfusion bath solution. The
middle current recordings were made 60 min after exposure of the oocyte
to dibutyryl-cAMP. Shortly after these currents were recorded,
dibutyryl-cAMP was removed from the perfusion bath solution; the
recordings at the bottom of the figure were made 30 min after removal
of dibutyryl-cAMP.